Zika virus (ZIKV) belongs to the genus of flavivirus within the family Flaviviridae. Many flaviviruses are significant human pathogens, including ZIKV, yellow fever (YFV), dengue virus (DENV serotypes 1 to 4), Japanese encephalitis virus (JEV), West Nile virus (WNV), and tick-borne encephalitis virus (TBEV). ZIKV is predominantly transmitted by Aedes spp. mosquitoes, which also transmit DENV and YFV, as well as chikungunya virus (an emerging alphavirus). Besides mosquitoes, ZIKV can also be transmitted through maternofetal route, sex, blood transfusion, and organ transplantation. Approximately 80% of the ZIKV infections are asymptomatic. Disease symptoms associated with ZIKV infection include headaches, fever, lethargy, rash, conjunctivitis, myalgia, and arthralgia. Severe diseases of ZIKV infection include neurotropic Guillain-Barre syndrome and congenital microcephaly. The flavivirus genome is a single-strand, positive-sense RNA of approximately 11,000 nucleotides. It contains a 5′ untranslated region (UTR), an open-reading frame (ORF), and a 3′ UTR. The single ORF encodes a long polyprotein which is processed into ten viral proteins, including three structural proteins—capsid (C), precursor membrane (prM), and envelope (E)—and seven non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5).
Diagnosis of ZIKV infection is performed through detection of viral components (e.g., viral RNA, viral proteins, or virus isolation) and detection of host immune response (e.g., antibodies against viral proteins). For viral component-based diagnosis, RT-PCR, immunoassay, and virus isolation detect ZIKV RNA, viral proteins, and live virus, respectively (Lanciotti et al., 2008); among them, RT-PCR is the most popular assay because of its sensitivity and specificity. The viremic phase of ZIKV infection usually lasts for about one week, yet occasionally persists beyond two weeks. Due to the short duration of the viremic phase, the diagnostic window for detection of viral components is narrow. Therefore, host immune response-based assays play an important role, among which enzyme-linked immunosorbent assays (ELISA), such as IgM-capture ELISA (MAC-ELISA), and plaque reduction neutralization test (PRNT) are the two most commonly used serologic assays in ZIKV diagnosis. Conventionally, serologic diagnosis of Zika virus (ZIKV) infection relies mainly upon IgM-capture ELISA which is confounded with the flaw of cross-reactivity among different flaviviruses. Unfortunately, the interpretation of conventional IgM-capture ELISA assays for ZIKV and other flaviviruses are challenging due to the cross-reactive nature of anti-flaviviral antibodies conventionally used in such tests, leading to equivocal diagnostic results. This challenge is confounding Zika diagnosis because (i) many flaviviruses (e.g., ZIKV and DENV) produce similar disease symptoms and (ii) antibodies from patients infected with ZIKV cross-react with other flaviviruses. Consequently, ZIKV IgM-capture ELISA results typically require neutralization tests for confirmation. Furthermore, PRNT is time-consuming, labor-intensive, slow, and low-throughput, and cost-ineffective, impairing attempts at rapid diagnosis to halt or slow spread if infection. Moreover, PRNT still relies upon both virus-specific and cross-reactive epitopes of viral E protein such that the results may be inconclusive with respect to flavivirus infections (Shan et al., 2016a). There is therefore a need to improve the accuracy and speed of serologic diagnosis for flaviviruses, ZIKV in particular.